Postreplicative recruitment of cohesin to double-strand breaks is required for DNA repair

Chromosome stability depends on accurate chromosome segregation and efficient DNA double-strand break (DSB) repair. Sister chromatid cohesion, established during S phase by the protein complex cohesin, is central to both processes. In the absence of cohesion, chromosomes missegregate and G2-phase DSB repair fails. Here, we demonstrate that G2-phase repair also requires the presence of cohesin at the damage site. Cohesin components are shown to be recruited to extended chromosome regions surrounding DNA breaks induced during G2. We find that in the absence of functional cohesin-loading proteins (Scc2/Scc4), the accumulation of cohesin at DSBs is abolished and repair is defective, even though sister chromatids are connected by S phase generated cohesion. Evidence is also provided that DSB induction elicits establishment of sister chromatid cohesion in G2, implicating that damage-recruited cohesin facilitates DNA repair by tethering chromatids.     

Mineral and carbon usage of two synthetic pyrethroid degrading bacterial isolates

AIMS: To investigate the biodegrading ability and cometabolism of synthetic pyrethroid (SP) utilizing bacteria in cultures with various minerals and carbon sources. METHODS AND RESULTS: Previously isolated SP-degrading Pseudomonas sp. and Serratia sp. were used in cultures containing either flumethrin SP or cypermethrin SP formulations. The culture media consisted of either (i) water only, (ii) water and sucrose, (iii) mineral broth or (iv) mineral broth and sucrose. The growth of both organisms was greatest in the mineral broth and sucrose medium, but the growth-limiting factor for Pseudomonas sp. strain Circle was the mineral content whereas for Serratia sp. strain White it was the carbon substrate. CONCLUSION: The greatest extent of degradation of both SP-based compounds occurred with Pseudomonas sp. strain Circle but was dependant on the medium. SIGNIFICANCE AND IMPACT OF THE STUDY: This investigation could lead to the development of a relatively inexpensive medium supplement to enhance the microbial biodegradation of undesirable compounds, either in situ or ex situ. In this particular case, for the biodegradation of SPs used in sheep dip.     

The interdigitated beta-helix domain of the P22 tailspike protein acts as a molecular clamp in trimer stabilization

The P22 tailspike adhesin is an elongated thermostable trimer resistant to protease digestion and to denaturation in sodium dodecyl sulfate. Monomeric, dimeric, and protrimeric folding and assembly intermediates lack this stability and are thermolabile. In the native trimer, three right-handed parallel beta-helices (residues 143-540), pack side-by-side around the three-fold axis. After residue 540, these single chain beta-helices terminate and residues 541-567 of the three polypeptide chains wrap around each other to form a three-stranded interdigitated beta-helix. Three mutants located in this region -- G546D, R563Q, and A575T -- blocked formation of native tailspike trimers, and accumulated soluble forms of the mutant polypeptide chains within cells. The substitutions R563Q and A575T appeared to prevent stable association of partially folded monomers. G546D, in the interdigitated region of the chain, blocked tailspike folding at the transition from the partially-folded protrimer to the native trimer. The protrimer-like species accumulating in the G546D mutant melted out at 42 degrees C and was trypsin and SDS sensitive. The G546D defect was not corrected by introduction of global suppressor mutations, which correct kinetic defects in beta-helix folding. The simplest interpretation of these results is that the very high thermostability (T(m) = 88 degrees C), protease and detergent resistance of the native tailspike acquired in the protrimer-to-trimer transition, depends on the formation of the three-stranded interdigitated region. This interdigitated beta-helix appears to function as a molecular clamp insuring thermostable subunit association in the native trimer.     

Genomic structure, chromosomal localization, and expression pattern of the human LIM-homeobox3 (LHX 3) gene

Lhx3 is a LIM-homeobox protein essential for pituitary development in mice. The human homologue gene spans 7.2 kb and contains 7 exons, including two alternatively spliced first exons. This structure encodes two distinct protein isoforms, LHX3a and LHX3b, that differ exclusively in their amino-terminus. The LHX3 gene was localized at 9q34.2-34.3. The predicted protein sequence is highly homologous to other known Lhx3 proteins, the highest degree of homology being in the conserved domains. The highest expression of LHX3 was found in pituitary gland, spinal cord, and lung. Among different pituitary cell types, corticotrophs appear to express preferentially LHX3b isoform, suggesting a distinct role of the b-form in the development of this cell lineage. Although the human LHX3 gene structure would provide a ground for clarification of the molecular basis of complete anterior pituitary deficiency, we were unable to identify any mutation in the LHX3 gene of 46 such patients.     

Role of rhamnolipid biosurfactants in the uptake and mineralization of hexadecane in Pseudomonas aeruginosa

A study was undertaken to investigate the mechanisms for biosurfactant-enhanced hexadecane uptake into Pseudomonas aeruginosa. Two strains of Ps. aeruginosa were studied, one producing rhamnolipids (PG201) and the other rhamnolipid deficient (UO299). Rhamnolipids produced by PG201 acted to increase the solubility of n-hexadecane in the culture medium (from 1.84 to 22.76 microg l(-1). Rates of(l4)C-n-hexadecane uptake and mineralization were higher in PG201 than in UO299. However, the degree of difference was lower than expected. Additional studies were carried out on the cell surface properties of the two strains. During growth on n-hexadecane, the cell surface hydrophobicity of both PG201 (50.5%) and UO299 (33.7%) increased compared with that observed in water-soluble growth substrates (7-8%). Studies were also carried out to ascertain any energy requirements for the transport of n-hexadecane into Ps. aeruginosa cells. The addition of CCCP (an inhibitor of cytochrome oxidase which thereby blocks oxidative phosphorylation) at a range of concentrations caused a marked decrease in n-hexadecane uptake, indicating that n-hexadecane uptake in Ps. aeruginosa is an energy-dependent process. These studies support the hypothesis of alkane transport into microbial cells by direct contact with larger alkane droplets and by pseudosolubilization. Also, it appears that both mechanisms occur simultaneously.     

Vaccination of macaques against pathogenic simian immunodeficiency virus with Venezuelan equine encephalitis virus replicon particles

Vaccine vectors derived from Venezuelan equine encephalitis virus (VEE) that expressed simian immunodeficiency virus (SIV) immunogens were tested in rhesus macaques as part of the effort to design a safe and effective vaccine for human immunodeficiency virus. Immunization with VEE replicon particles induced both humoral and cellular immune responses. Four of four vaccinated animals were protected against disease for at least 16 months following intravenous challenge with a pathogenic SIV swarm, while two of four controls required euthanasia at 10 and 11 weeks. Vaccination reduced the mean peak viral load 100-fold. The plasma viral load was reduced to below the limit of detection (1,500 genome copies/ml) in one vaccinated animal between 6 and 16 weeks postchallenge and in another from week 6 through the last sampling time (40 weeks postchallenge). The extent of reduction in challenge virus replication was directly correlated with the strength of the immune response induced by the vectors, which suggests that vaccination was effective.     

Effect of protein hydrolysate on the degradation of diesel fuel in soil*

The addition of protein hydrolysate solution to soil contaminated with diesel fuel was investigated for effects on diesel degradation. The application of protein hydrolysate solution led to an increase in the removal of diesel from the soil. At the end of the 21d experimental period the amount of diesel removed from the soil was 21% greater with the addition of protein hydrolysate solution when compared to a control system. This increased removal was linked to increases in both the number of hydrocarbon degrading micro-organisms, and an extension to their period of activity.